REGULATORY MECHANISMS IN THE IMMUNE SYSTEM

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The overall goal of this research program is to understand the affinities/avidities of the sites of interaction on T cell ab receptors (TCRs) for conventional peptide antigens or superantigens (SAgs) bound to major histocompatibility complex (MHC) Class II molecules and how the quality of that interaction affects the fate of the T cell in positive selection, negative selection or productive activation. Our experiments will be performed in vitro with recombinant soluble proteins. We will use mutational studies to determine in detail the sites of interaction of three staphylococcal SAgs, SEA, SEC3 and TSST-1 on mouse Vbeta3, Vbeta8.2 and Vbeta15 respectively. In addition, in biosensor surface plasmon resonance studies we will examine the contribution from Va and MHC class II to the quality of TCR Vbeta binding to these SAgs. In a separate set of experiments, we will prepare soluble TCRs from CD4 plus T cell hybridomas of known antigen/MHC reactivity derived from T cells that have been positively selected in transgenic mice bearing a single MHC class II molecule complexed to a single peptide. We will use these TCRs to compare the affinities/avidities required for positive selection, negative selection and T cell activation. The experiments will involve direct binding studies of soluble TCRs to MHC/peptide complexes and include construction of soluble multivalent TCRs with defined geometries. These experiments will test hypotheses which propose that TCR affinity determines the fate of developing T cells.